P410: LPIN1 REGULATES LIPID HOMEOSTASIS AND MAINTAINS STEM CELL ENRICHED POPULATIONS IN AML
Authors: K. Huber, S. Garg, L. He, A. Pouya, C. Rohde, C. Lüchtenborg, C. Arnold, J. Zaugg, S. Raffel, C. Müller-Tidow, B. Brügger, C. Pabst
Journal: HemaSphere (2022)

Background: Leukemic stem cells (LSCs) often escape chemotherapy treatment due to the acquisition of stem cell like properties such as slow cell cycle progression, which makes them main drivers of cytologic relapse in acute myeloid leukemia (AML). Identification of novel therapeutic targets to specifically eradicate residual chemoresistant AML cells therefore represents a key challenge in AML therapy. Since LSCs show altered metabolic features, which include dependency on low ROS levels and oxidative phosphorylation of amino acids, targeting LSC metabolism has evolved as a promising approach. Recently the phospholipid phosphatidylserine was shown to be required for LSC stemness and differentiation. However, the role of the main phospholipid synthesis pathway in which the phosphatidic acid phosphatase (PAP) lipin1 catalyzes a rate limiting step remains largely unexplored. Aims: We investigated the role of the phosphatidic acid phosphatase LPIN1 in AML and healthy hematopoietic cells. Methods: We applied a gene depletion approach using shRNAs and CRISPR/Cas9 on AML cell lines, primary AML samples and cord blood derived CD34+ hematopoietic stem and progenitor cells (HSPCs) to monitor the effects of LPIN1 depletion on proliferation, differentiation, colony forming capacity and bone marrow engraftment in mice. To gain insights into the mechanisms of action, we combined RNA-sequencing (RNA-seq) with lipidomics assays. Results: Here we show that regulation of lipid homeostasis by lipin1 is required for HSPC and LSC function. RNA-seq data analysis of healthy hematopoietic cells revealed highest LPIN1 expression in the immature CD34 positive compartment, which steadily decreases during myeloid differentiation. In normal karyotype (NK) AML LPIN1 was utmost expressed in an AML subpopulation characterized by high LSC frequency and co-positivity for the LSC markers CD34 and GPR56. Knock down (KD) of LPIN1 in CD34 positive HSPCs and patient derived xenograft (PDX) leukemic cells hampered proliferation and reduced the CD34 positive LSC compartment in vitro. In xenotransplantation assays LPIN1 KD reduced bone marrow engraftment of healthy HSPC and human AML cells. Furthermore, in vivo differentiation of CD34 positive cells into B-cells was almost completely abrogated upon LPIN1 KD. In the leukemia setting, LPIN1 KD particularly reduced the LSC-enriched CD34+GPR56+ co-positive fraction. While RNA-seq of the AML cell line OCI-AML3 and CD34 positive cells showed only subtle gene expression changes in phospholipid metabolism upon LPIN1 KD, lipidomics analyses revealed significant differences in the composition of phospholipids upon LPIN1 KD. Summary/Conclusion: Our data establish LPIN1 as an essential enzyme maintaining lipid homeostasis and stem cell functions in the hematopoietic system. Identification of phospholipid entities with differential impact on healthy versus leukemic cells might help refine LSC targeted therapies.